Circadian rhythms are endogenous rhythms that help keep organisms in synchrony with their environment and contribute to an organism's fitness for life on earth. The importance of circadian rhythms has been shown for many different health issues, from the timing of cancer treatments (Gorbacheva et al., 2005) to obesity and metabolic syndrome (Turek et al., 2005). Understanding the molecular basis of these rhythms will help us find ways to affect the processes they regulate in humans. Our long term goal is to understand in detail how the circadian clock functions in the model system Arabidopsis, and how both transcriptional and post-transcriptional processes are used to give this endogenous timekeeper a period of about a day in constant conditions of light and temperature. We will focus particularly on the putative central oscillator component CCA1 and extend our studies with CCA1 to other putative components such as LHY. Our specific experimental aims are: 1. To understand how CCA1 and LHY function in generating circadian rhythms, a. To test whether they (and other proposed components) are truly central oscillator components by testing whether a pulse of the protein can reset the phase of circadian rhythms, b. To test what allows rhythms to continue for a short time when CCA1 is absent and LHY is reduced and whether a line that is completely null for both CCA1 and LHY stops all rhythms immediately, c. To understand how the transcription of CCA1 is regulated, d. To identify the genome wide targets of CCA1 and LHY. 2. To understand the roles of post-transcriptional processes in maintaining circadian rhythms, a. To understand the roles that phosphorylation of CCA1 and LHY, particularly by the protein kinase CK2, plays in their functions, b. To understand how degradation of the CCA1 and LHY proteins (which themselves cycle with a circadian rhythm) is regulated, c. To see whether CCA1-LHY protein interactions and localization dynamics in the cell play a role in circadian regulation. 3. To identify and characterize additional genes/proteins that are involved with the circadian clock, a. To understand the role of our recently discovered CCA1-interacting protein (designated CIRP), which affects the period of circadian rhythms, b. To investigate the possible significance of CCA1 interaction with histone H3. c. To identify the genes corresponding to the mutants that we have isolated that have altered circadian rhythms and to develop an understanding of the roles of these genes. [unreadable] [unreadable] [unreadable]